LOCUS       sequence-233069-        7126 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of the human Rad18 Ubiquitin binding domain and
            BFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7126)
  AUTHORS   Bashir S, Dang T, Rossius J, Wolf J, Kuhn R
  TITLE     Enhancement of CRISPR-Cas9 induced precise gene editing by targeting
            histone H2A-K15 ubiquitination.
  JOURNAL   BMC Biotechnol. 2020 Oct 23;20(1):57. doi: 
            10.1186/s12896-020-00650-x.
  PUBMED    33097066
REFERENCE   2  (bases 1 to 7126)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "BMC 
            Biotechnol."; date: "2020-10-23"; volume: "20(1):57. doi"; pages: " 
            10.1186/s12896-020-00650-x"
COMMENT     Sequence Label: pCAG-Rad18UBD
FEATURES             Location/Qualifiers
     source          1..7126
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     2..18
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2..18
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     enhancer        38..417
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          697..1713
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1637..1659
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1726..1745
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1726..1744
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1746..1755
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1755..1778
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(1940..1956)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    1976..2200
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     intron          2421..3359
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3316..3336
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      3394..3403
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3400..3402,3403..3405,3406..4119)
                     /codon_start=1
                     /product="enhanced blue variant of GFP (Ai et al., 2007)"
                     /label=EBFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSHGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGVDFKEDGNILGHKLEYNYNSHNIYIMADKQKNGIK
                     VNFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDSHYLSTQSVLSKDPNEKRDHMVLL
                     EFRTAAGITLGMDELYK"
     primer_bind     complement(3445..3466)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     polyA_signal    4150..4261
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4379..4396)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4379..4395)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4388..4410)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      complement(4536..4991)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4673..4694)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4885..4904
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5018..5122
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5123..5191,5192..5983)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5341..5360)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6154..6742
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6643..6662
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6896..6913
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7030..7051
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7066..7071,7072..7089,7090..7096)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7104..7120
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     join(7109..7126,1..5)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
ORIGIN
        1 acaggaaaca gctatgacca tgaggcgcgc cggattcgac attgattatt gactagttat
       61 taatagtaat caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca
      121 taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca
      181 ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg
      241 gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg
      301 ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc
      361 ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtc
      421 gaggtgagcc ccacgttctg cttcactctc cccatctccc ccccctcccc acccccaatt
      481 ttgtatttat ttatttttta attattttgt gcagcgatgg gggcgggggg gggggggggg
      541 cgcgcgccag gcgggggggg gggggggggg gggggggggg gggggggggg ggggggggcg
      601 gcggcagcca atcagagcgg cgcgctccga aagtttcctt ttatggcgag gcggcggcgg
      661 cggcggccct ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgcg ctgccttcgc
      721 cccgtgcccc gctccgccgc cgcctcgcgc cgcccgcccc ggctctgact gaccgcgtta
      781 ctcccacagg tgagcgggcg ggacggccct tctcctccgg gctgtaatta gcgcttggtt
      841 taatgacggc ttgtttcttt tctgtggctg cgtgaaagcc ttgaggggct ccgggagggc
      901 cctttgtgcg gggggagcgg ctcggggggt gcgtgcgtgt gtgtgtgcgt ggggagcgcc
      961 gcgtgcggct ccgcgctgcc cggcggctgt gagcgctgcg ggcgcggcgc ggggctttgt
     1021 gcgctccgca gtgtgcgcga ggggagcgcg gccgggggcg gtgccccgcg gtgcgggggg
     1081 ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg tgggggggtg agcagggggt
     1141 gtgggcgcgt cggtcgggct gcaacccccc ctgcaccccc ctccccgagt tgctgagcac
     1201 ggcccggctt cgggtgcggg gctccgtacg gggcgtggcg cggggctcgc cgtgccgggc
     1261 ggggggtggc ggcaggtggg ggtgccgggc ggggcggggc cgcctcgggc cggggggggc
     1321 tcgggggggg ggcgcggcgg cccccggagc gccggcggct gtcgaggcgc ggcgagccgc
     1381 agccattgcc ttttatggta atcgtgcgag agggcgcagg gacttccttt gtcccaaatc
     1441 tgtgcggagc cgaaatctgg gaggcgccgc cgcaccccct ctagcgggcg cggggcgaag
     1501 cggtgcggcg ccggcaggaa ggaaatgggc ggggagggcc ttcgtgcgtc gccgcgccgc
     1561 cgtccccttc tccctctcca gcctcggggc tgtccgcggg gggacggctg ccttcggggg
     1621 ggacggggca gggcggggtt cggcttctgg cgtgtgaccg gcggctctag agcctctgct
     1681 aaccatgttc atgccttctt ctttttccta cagatcctta attaataata cgactcacta
     1741 taggggccac catggactac aaggatgacg acgataaacc ctcgacatcc actttgaaac
     1801 aagttactaa agtggattgt cctgtttgcg gggttaacat tccagaaagt aagcattttg
     1861 acagctgttt atcacgcgaa gagaagaagg aaagcctcag aagttctgtt cacaaaaggt
     1921 agacgcgtaa atgattgcag atccactagt tctagagctc gctgatcagc ctcgactgtg
     1981 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     2041 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt
     2101 aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa
     2161 gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc
     2221 agctggggct cgagatccac tagttctagc ctcgaggcta gagcggcccc ggtgcctaga
     2281 gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta ctggctccgc ctttttcccg
     2341 agggtggggg agaaccgtat ataagtgcag tagtcgccgt gaacgttctt tttcgcaacg
     2401 ggtttgccgc cagaacacag gtaagtgccg tgtgtggttc ccgcgggcct ggcctcttta
     2461 cgggttatgg cccttgcgtg ccttgaatta cttccacctg gctgcagtac gtgattcttg
     2521 atcccgagct tcgggttgga agtgggtggg agagttcgag gccttgcgct taaggagccc
     2581 cttcgcctcg tgcttgagtt gaggcctggc ctgggcgctg gggccgccgc gtgcgaatct
     2641 ggtggcacct tcgcgcctgt ctcgctgctt tcgataagtc tctagccatt taaaattttt
     2701 gatgacctgc tgcgacgctt tttttctggc aagatagtct tgtaaatgcg ggccaagatc
     2761 tgcacactgg tatttcggtt tttggggccg cgggcggcga cggggcccgt gcgtcccagc
     2821 gcacatgttc ggcgaggcgg ggcctgcgag cgcggccacc gagaatcgga cgggggtagt
     2881 ctcaagctgg ccggcctgct ctggtgcctg gcctcgcgcc gccgtgtatc gccccgccct
     2941 gggcggcaag gctggcccgg tcggcaccag ttgcgtgagc ggaaagatgg ccgcttcccg
     3001 gccctgctgc agggagctca aaatggagga cgcggcgctc gggagagcgg gcgggtgagt
     3061 cacccacaca aaggaaaagg gcctttccgt cctcagccgt cgcttcatgt gactccacgg
     3121 agtaccgggc gccgtccagg cacctcgatt agttctcgag cttttggagt acgtcgtctt
     3181 taggttgggg ggaggggttt tatgcgatgg agtttcccca cactgagtgg gtggagactg
     3241 aagttaggcc agcttggcac ttgatgtaat tctccttgga atttgccctt tttgagtttg
     3301 gatcttggtt cattctcaag cctcagacag tggttcaaag tttttttctt ccatttcagg
     3361 tgtcgtgagc gatcgcatcg acagtgttta aacgccacca tggtgagcaa gggcgaggag
     3421 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
     3481 ttcagcgtga gcggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc
     3541 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgagccac
     3601 ggcgtgcagt gcttcgcccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
     3661 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcacctac
     3721 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     3781 ggcgtcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac
     3841 agccacaaca tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag
     3901 atccgccaca acgtggagga cggcagcgtg cagctcgccg accactacca gcagaacacc
     3961 cccatcggcg acggccccgt gctgctgccc gacagccact acctgagcac ccagtccgtg
     4021 ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt ccgcaccgcc
     4081 gccgggatca ctctcggcat ggacgagctg tacaagtgag aattcctaga gctcgctgat
     4141 cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt
     4201 ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat
     4261 cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg
     4321 gggaggattg ggaagagaat agcaggcatg cgtttaaaca ttagtcgaca gggccgccac
     4381 tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc
     4441 ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc
     4501 cttcccaaca gttgcgcagc ctgaatggcg aatggaaatt gtaagcgtta atattttgtt
     4561 aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg
     4621 caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg
     4681 gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta
     4741 tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg
     4801 ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa
     4861 gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct
     4921 ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct
     4981 acagggcgcg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     5041 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
     5101 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
     5161 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     5221 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
     5281 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
     5341 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
     5401 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
     5461 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     5521 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
     5581 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     5641 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
     5701 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
     5761 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     5821 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
     5881 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     5941 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
     6001 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     6061 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     6121 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     6181 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     6241 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     6301 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     6361 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     6421 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     6481 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     6541 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     6601 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     6661 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     6721 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     6781 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     6841 tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga
     6901 gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg
     6961 gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg
     7021 caacgcaatt aatgtgagtt agctcactca ttaggcaccc caggctttac actttatgct
     7081 tccggctcgt atgttgtgtg gaattgtgag cggataacaa tttcac
//
